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1.
Curr Opin Genet Dev ; 63: 30-35, 2020 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-32172182

RESUMO

Study of early human embryo development is essential for advancing reproductive and regenerative medicine. Traditional human embryological studies rely on embryonic tissue specimens, which are difficult to acquire due to technical challenges and ethical restrictions. The availability of human stem cells with developmental potentials comparable to pre-implantation and peri-implantation human embryonic and extraembryonic cells, together with properly engineered in vitro culture environments, allow for the first time researchers to generate self-organized multicellular structures in vitro that mimic the structural and molecular features of their in vivo counterparts. The development of these stem cell-based, synthetic human embryo models offers a paradigm-shifting experimental system for quantitative measurements and perturbations of multicellular development, critical for advancing human embryology and reproductive and regenerative medicine without using intact human embryos.

2.
Small ; : e1907521, 2020 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-32174029

RESUMO

The development of a powerful immunoassay platform with capacities of both simplicity and high multiplexing is promising for disease diagnosis. To meet this urgent need, for the first time, a multiplexed luminescent oxygen channeling immunoassay (multi-LOCI) platform by implementation of LOCI with suspension array technology is reported. As the microcarrier of the platform, a unique dual-functional barcode with a host-guest structure composed of a quantum dot host bead (QDH) and LOCI acceptor beads (ABs) is designed, in which QDH provides function of high coding capacity while ABs facilitate the LOCI function. The analytes bridge QDH@ABs and LOCI donor beads (DBs) into a close proximity, forming a QDH@ABs-DBs "host-guest-satellite" superstructure that generates both barcode signal from QDH and LOCI signal induced by singlet oxygen channeling between ABs and DBs. Through imaging-based decoding, different barcodes are automatically distinguished and colocalized with LOCI signals. Importantly, the assay achieves simultaneous detection of multiple analytes within one reaction, simply by following a "mix-and-measure" protocol without the need for tedious washing steps. Furthermore, the multi-LOCI platform is validated for real sample measurements. With the advantages of robustness, simplicity, and high multiplexing, the platform holds great potential for the development of point-of-care diagnostics.

3.
Cell Rep ; 29(13): 4568-4582.e5, 2019 Dec 24.
Artigo em Inglês | MEDLINE | ID: mdl-31875561

RESUMO

In vitro gametogenesis is the process of making germline cells from human pluripotent stem cells. The foundation of this model is the quality of the first progenitors called primordial germ cells (PGCs), which in vivo are specified during the peri-implantation window of human development. Here, we show that human PGC (hPGC) specification begins at day 12 post-fertilization. Using single-cell RNA sequencing of hPGC-like cells (hPGCLCs) differentiated from pluripotent stem cells, we discovered that hPGCLC specification involves resetting pluripotency toward a transitional state with shared characteristics between naive and primed pluripotency, followed by differentiation into lineage-primed TFAP2A+ progenitors. Applying the germline trajectory to TFAP2C mutants reveals that TFAP2C functions in the TFAP2A+ progenitors upstream of PRDM1 to regulate the expression of SOX17. This serves to protect hPGCLCs from crossing the Weismann's barrier to adopt somatic cell fates and, therefore, is an essential mechanism for successfully initiating in vitro gametogenesis.

4.
Nature ; 573(7774): 421-425, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31511693

RESUMO

Early human embryonic development involves extensive lineage diversification, cell-fate specification and tissue patterning1. Despite its basic and clinical importance, early human embryonic development remains relatively unexplained owing to interspecies divergence2,3 and limited accessibility to human embryo samples. Here we report that human pluripotent stem cells (hPSCs) in a microfluidic device recapitulate, in a highly controllable and scalable fashion, landmarks of the development of the epiblast and amniotic ectoderm parts of the conceptus, including lumenogenesis of the epiblast and the resultant pro-amniotic cavity, formation of a bipolar embryonic sac, and specification of primordial germ cells and primitive streak cells. We further show that amniotic ectoderm-like cells function as a signalling centre to trigger the onset of gastrulation-like events in hPSCs. Given its controllability and scalability, the microfluidic model provides a powerful experimental system to advance knowledge of human embryology and reproduction. This model could assist in the rational design of differentiation protocols of hPSCs for disease modelling and cell therapy, and in high-throughput drug and toxicity screens to prevent pregnancy failure and birth defects.

5.
Lab Chip ; 19(18): 3065-3076, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31389447

RESUMO

Widespread commercial and clinical adaptation of biomedical microfluidic technology has been limited in large part due to the lack of mass producibility of polydimethylsiloxane (PDMS) and glass-based devices commonly as reported in the literature. Here, we present a batch-fabricated, robust, and mass-producible immunophenotyping microfluidic device using silicon micromachining processes. Our Si and glass-based microfluidic device, named the silicon microfluidic immunophenotyping assay (SiMIPA), consists of a highly porous (∼40%) silicon membrane that can selectively separate microparticles below a certain size threshold. The device is capable of isolating and stimulating specific leukocyte populations, and allows for measuring their secretion of cell signaling proteins by means of a no-wash homogeneous chemiluminescence-based immunoassay. The high manufacturing throughput (∼170 devices per wafer) makes a large quantity of SiMIPA chips readily available for clinically relevant applications, which normally require large dataset acquisitions for statistical accuracy. With 30 SiMIPA chips, we performed in vitro immunomodulatory drug screening on isolated leukocyte subsets, yielding 5 data points at 6 drug concentrations. Furthermore, the excellent structural integrity of the device allowed for samples and reagents to be loaded using a micropipette, greatly simplifying the experimental protocol.

6.
Front Genet ; 10: 603, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31312211

RESUMO

Circular RNAs (circRNAs) were recently recognized to act as competing endogenous RNAs and play roles in gene expression regulation. Previous studies in humans and silkworms have shown that circRNAs take part in immune regulation. Here, we conducted coelomocyte circRNA sequencing to explore its immune functions in healthy and skin ulceration syndrome (SUS)-diseased sea cucumbers. A total of 3,592 circRNAs were identified in libraries with diversified circularization patterns compared with animal models. The common intron-pairing-driven circularization models are not popular in sea cucumber genome, which was replaced with intergenic region circularization. The accuracy of these identified circRNAs was further validated by Sanger sequencing and RNase R-treated assays. Expression profile analysis indicated that 117 circRNAs were upregulated and 144 circRNAs were downregulated in SUS-diseased condition, of which 71.6% were intergenic-type circRNAs. The interaction network of differentially expressed circRNAs and microRNAs (miRNAs) was constructed and showed that miR-2008 and miR-31, detected with significantly differential expression in SUS-affected samples in a previous study, were predicted to be regulated by 10 and 11 differentially expressed circRNAs with more than 10 binding sites, respectively. Moreover, seven circRNAs were further validated by quantitative real-time PCR, whose variation trends were consistent with circRNA sequencing. All our results supported that intergenic-type circRNAs might have a dominant function in Apostichopus japonicas immune response by acting as miRNA regulators.

7.
Biomaterials ; 216: 119244, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31207406

RESUMO

During early post-implantation human embryogenesis, the epiblast (EPI) within the blastocyst polarizes to generate a cyst with a central lumen. Cells at the uterine pole of the EPI cyst then undergo differentiation to form the amniotic ectoderm (AM), a tissue essential for further embryonic development. While the causes of early pregnancy failure are complex, improper lumenogenesis or amniogenesis of the EPI represent possible contributing factors. Here we report a novel AM microtissue array platform that allows quantitative phenotyping of lumenogenesis and amniogenesis of the EPI and demonstrate its potential application for embryonic toxicity profiling. Specifically, a human pluripotent stem cell (hPSC)-based amniogenic differentiation protocol was developed using a two-step micropatterning technique to generate a regular AM microtissue array with defined tissue sizes. A computer-assisted analysis pipeline was developed to automatically process imaging data and quantify morphological and biological features of AM microtissues. Analysis of the effects of cell density, cyst size and culture conditions revealed a clear connection between cyst size and amniogenesis of hPSC. Using this platform, we demonstrated that pharmacological inhibition of ROCK signaling, an essential mechanotransductive pathway, suppressed lumenogenesis but did not perturb amniogenic differentiation of hPSC, suggesting uncoupled regulatory mechanisms for AM morphogenesis vs. cytodifferentiation. The AM microtissue array was further applied to screen a panel of clinically relevant drugs, which successfully detected their differential teratogenecity. This work provides a technological platform for toxicological screening of clinically relevant drugs for their effects on lumenogenesis and amniogenesis during early human peri-implantation development, processes that have been previously inaccessible to study.

8.
Fish Shellfish Immunol ; 89: 727-735, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30981886

RESUMO

Caspase-8, an initiator caspase, plays a vital role in apoptosis. In this study, caspase-8-like (named as Cicaspase-8-like), a homologue of caspase-8, was identified in grass carp (Ctenopharygodon idella). The full-length cDNA sequence of CiCaspase-8-like was 1409 bp and contained a 162 bp 5'-UTR, a 239 bp 3'-UTR and a 1008 bp coding sequence. The putative amino acids sequence was 335 residues long, including a large subunit (P20) and a small subunit (P10), but lacking conserved death effector domains. A histidine active site DHSQMDAFVCCVLSHG and a cysteine active-site motif KPKLFFIQACQG were found in P20. Phylogenetic analysis showed that Cicaspase-8-like clustered with the caspase-8 and caspase-8-like of other fish and grouped closely with Carassius auratus caspase-8-like. Quantitative real-time PCR revealed that the Cicaspase-8-like mRNA were expressed constitutively in all tested tissues from healthy grass carp, with high expression level in the blood, spleen, liver and gill, indicating its role in immune reaction. The expression of Cicaspase-8-like mRNA was decreased significantly in the liver because of the stress caused by microcystin-LR (MC-LR) (75 and 100 µg MC-LR/kg BW) at 24 h and 96 h post injection (P < 0.05), but it was increased significantly in grass carp treated with 25 µg MC-LR/kg BW at 24 h (P < 0.05) post injection. Cleaved fragments of Cicaspase-8-like were observed using western blot analysis, and the expression of Cicaspase-8-like protein was increased after MC-LR treatments. Moreover, the expression of both caspase-9 and caspase-3 mRNA increased significantly after treatment with the three doses of MC-LR. TUNEL assay results showed remarkable changes in apoptosis after the MC-LR treatment. These results suggest that Cicaspase-8-like is an important caspase and plays an essential role in MC-LR-induced apoptosis.


Assuntos
Carpas/genética , Carpas/imunologia , Caspase 8/genética , Caspase 8/imunologia , Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Sequência de Aminoácidos , Animais , Caspase 8/química , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Microcistinas/efeitos adversos , Filogenia , Alinhamento de Sequência/veterinária
9.
Fish Shellfish Immunol ; 88: 403-406, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30862516

RESUMO

IFN-γ is a pleiotropic cytokine with significant roles in antiviral, antitumor and immune regulation. It could be used as an immuno-enhancer to improve fish protectiveness against pathogens. In this study, the prokaryotic expression plasmid pTwin1-N-IFN-γ was constructed to express Cyprinus carpio (common carp) IFN-γ fused with a chitin binding domain (CBD) and a self-cleavable intein-tag, Synechocystis sp DnaB. The recombinant protein CBD-DnaB-IFN-γ with the molecular weight of 44.25 kD was successfully expressed in soluble form, and the rIFN-γ (approximate 18.61 kD) was further cleaved and eluted under pH = 7.0 at 25 °C. rIFN-γ could be recognized by western blotting with rabbit anti-grass carp IFN-γ polyclonal antibody. Cytotoxicity studies on EPC cells showed that only 500 ng/ml rIFN-γ had a subtle effect on cells growth and its proliferation rate was reduced to 76.2%. EPC cells incubated with 100 ng/ml rIFN-γ showed significantly higher resistance against SVCV, reducing the TCID50/ml by more than 800-fold. In vivo studies suggested that intraperitoneal injection of rIFN-γ significantly improved the survival rate of common carps compared with SVCV challenge alone. These results implied that rIFN-γ would act as an immuno-enhancer in carp aquaculture.


Assuntos
Carpas/virologia , Doenças dos Peixes/prevenção & controle , Inteínas/genética , Interferon gama/imunologia , Infecções por Rhabdoviridae/veterinária , Animais , Aquicultura , Carpas/imunologia , Linhagem Celular Tumoral , Doenças dos Peixes/imunologia , Doenças dos Peixes/virologia , Imunidade Inata , Interferon gama/genética , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/imunologia , Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/prevenção & controle , Viremia/imunologia
10.
Rev Sci Instrum ; 90(2): 023107, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30831709

RESUMO

Nanoparticles are important in several areas of modern biomedical research. However, detection and characterization of nanoparticles is challenging due to their small size. Back-focal-plane interferometry (BFPI) is a highly sensitive technique that has been used in laser tweezers for quantitative measurement of force and displacement. The utility of BFPI for detection and characterization of nanoparticles, however, has not yet been achieved. Here we show that BFPI can be used for rapid probing of a suspension of nanoparticles in a spatially confined microfluidic channel. We show that the Gaussian Root-mean-squared noise of the BFPI signal is highly sensitive to the nanoparticle size and can be used as a parameter for rapid detection of nanoparticles at a single-particle level and characterization of particle heterogeneities in a suspension. By precisely aligning the optical trap relative to the channel boundaries, individual polystyrene particles with a diameter as small as 63 nm can be detected using BFPI with a high signal-to-noise ratio.

11.
Small ; 15(5): e1802891, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30632269

RESUMO

Cancer stem-like cells (CSCs) have been shown to initiate tumorigenesis and cancer metastasis in many cancer types. Although identification of CSCs through specific marker expression helps define the CSC compartment, it does not directly provide information on how or why this cancer cell subpopulation is more metastatic or tumorigenic. In this study, the functional and biophysical characteristics of aggressive and lethal inflammatory breast cancer (IBC) CSCs at the single-cell level are comprehensively profiled using multiple microengineered tools. Distinct functional (cell migration, growth, adhesion, invasion and self-renewal) and biophysical (cell deformability, adhesion strength and contractility) properties of ALDH+ SUM149 IBC CSCs are found as compared to their ALDH- non-CSC counterpart, providing biophysical insights into why CSCs has an enhanced propensity to metastasize. It is further shown that the cellular biophysical phenotype can predict and determine IBC cells' tumorigenic ability. SUM149 and SUM159 IBC cells selected and modulated through biophysical attributes-adhesion and stiffness-show characteristics of CSCs in vitro and enhance tumorigenicity in in vivo murine models of primary tumor growth. Overall, the multiparametric cellular biophysical phenotyping and modulation of IBC CSCs yields a new understanding of IBC's metastatic properties and how they might develop and be targeted for therapeutic interventions.

12.
Acta Biomater ; 86: 125-134, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30641291

RESUMO

Formation of the primitive streak (PS) marks one of the most important developmental milestones in embryonic development. However, our understanding of cellular mechanism(s) underlying cell fate diversification along the anterior-posterior axis of the PS remains incomplete. Furthermore, differences in biophysical phenotypes between anterior and posterior PS cells, which could affect their functions and regulate their fate decisions, remain uncharacterized. Herein, anterior and posterior PS cells were derived using human pluripotent stem cell (hPSC)-based in vitro culture systems. We observed that anterior and posterior PS cells displayed significantly different biophysical phenotypes, including cell morphology, migration, and traction force generation, which was further regulated by different levels of Activin A- and BMP4-mediated developmental signaling. Our data further suggested that intracellular cytoskeletal contraction could mediate anterior and posterior PS differentiation and phenotypic bifurcation through its effect on Activin A- and BMP4-mediated intracellular signaling events. Together, our data provide new information about biophysical phenotypes of anterior and posterior PS cells and reveal an important role of intracellular cytoskeletal contractility in regulating anterior and posterior PS differentiation of hPSCs. STATEMENT OF SIGNIFICANCE: Formation of the primitive streak (PS) marks one of the most important developmental milestones in embryonic development. However, molecular and cellular mechanism(s) underlying functional diversification of embryonic cells along the anterior-posterior axis of the PS remains incompletely understood. This work describes the first study to characterize the biophysical properties of anterior and posterior PS cells derived from human pluripotent stem cells (hPSCs). Importantly, our data showing the important role of cytoskeleton contraction in controlling anterior vs. posterior PS cell phenotypic switch (through its effect on intracellular Smad signaling activities downstream of Activin A and BMP4) should shed new light on biomechanical regulations of the development and anterior-posterior patterning of the PS. Our work will contribute significantly to uncovering new biophysical principles and cellular mechanisms driving cell lineage diversification and patterning during the PS formation.

13.
Int J Biol Macromol ; 121: 443-453, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30316769

RESUMO

P-glycoprotein (P-gp or ABCB1) and multidrug resistance associated proteins (MRPs or ABCC) belonging to the family of ATP-binding cassette (ABC) transporters protect aquatic organisms from various toxicants and pathogen exposure. The function of ABC transporters of Sinonovacula constricta in response to pathogens was explored by cloning the complete cDNA of ABCB1 and ABCC5 of S. constricta through the rapid amplification of cDNA ends. Tissue-specific expression showed that ABCB1 and ABCC5 have the highest expression in hemocytes and gills among eight examined tissues, respectively. The transport activities of ABCB1 and ABCC5 in the gills were severely inhibited after their corresponding inhibitor treatments. The expression levels and transport activities of ABCB1 and ABCB5 were also investigated by using S. constricta samples infected by Vibrio parahaemolyticus at a final concentration of 107 CFU/mL. Results showed that the mRNA expression levels of ABCB1 and ABCC5 were significantly down-regulated when exposed to V. parahaemolyticus at 6 h and 12 h (P < 0.05) and then recovered to normal level at 24 h and 48 h. Consistently, the activities of the two ABC transporters display the same trends as the mRNA expression levels. The results, combined with that of previous investigations, indicated that these activities may be an innate immune response in S. constricta. Immunohistochemical results justified the tissue localization of P-gp in the interface between external environment and tissues. Taken together, our findings demonstrated that ABC transporters form an active, physiological barrier at the tissue-environment interface in S. constricta gills in response to pathogen infection, and they might play a vital role in adaptation and response to environmental stress and the immune defense.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Bivalves/microbiologia , Brânquias/metabolismo , Vibrio parahaemolyticus/fisiologia , Transportadores de Cassetes de Ligação de ATP/química , Sequência de Aminoácidos , Animais , Bivalves/imunologia , Bivalves/metabolismo , Regulação da Expressão Gênica , Imunidade Inata , Transporte Proteico
14.
Dev Comp Immunol ; 90: 55-59, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30172908

RESUMO

The reduction of disulfide bonds of exogenous antigens is crucial to the MHC-II class antigen processing and presenting pathway and is catalysed by interferon-γ-inducible lysosomal thiol reductase (GILT). In this study, a reptile GILT gene from Chinese soft-shelled turtle, Pelodiscus sinensis (PsGILT), was identified. The full-length cDNA of PsGILT is 1631 nucleotides (nt), including a 5'-untranslated region (UTR) of 3 nt, a 3'-UTR of 860 nt and an open reading frame (ORF) of 768 nt encoding 255 amino acids (aa). The conserved features in known GILTs, such as signal peptide, CXXC motif, GILT signature sequence, N-glycosylation site and conserved cysteines, were all found in the putative PsGILT protein. Genomic analysis revealed that PsGILT kept the "7 exons and 6 introns" structure of vertebrate GILT genes. PsGILT was expressed in all examined organs/tissues and was mainly expressed in spleen and blood. Increased mRNA expression levels of PsIFN-γ and PsGILT in PBLs were observed after induction with LPS, PolyI:C and recombinant IFN-γ (rIFN-γ). We also tested the reductase activity of rGILT in vitro and found that it could reduce intact human IgG into H chains and L chains. These above results implied that PsGILT may play an important role in resisting bacterial and viral infections, like other vertebrate GILTs.


Assuntos
Leucócitos Mononucleares/fisiologia , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/genética , Proteínas de Répteis/genética , Baço/fisiologia , Tartarugas/imunologia , Animais , Antivirais/metabolismo , Células Cultivadas , Clonagem Molecular , Humanos , Imunidade Inata , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/metabolismo , Filogenia , Poli I-C/imunologia , Proteínas de Répteis/metabolismo , Transcriptoma
16.
Small ; 14(50): e1803137, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30427572

RESUMO

Early human embryogenesis is a dynamic developmental process, involving continuous and concomitant changes in gene expression, structural reorganization, and cellular mechanics. However, the lack of investigation methods has limited the understanding of how cellular mechanical properties change during early human embryogenesis. In this study, ultrasound actuation of functionalized microbubbles targeted to integrin (acoustic tweezing cytometry, ATC) is employed for in situ measurement of cell stiffness during human embryonic stem cell (hESC) differentiation and morphogenesis. Cell stiffness, which is regulated by cytoskeleton structure, remains unchanged in undifferentiated hESCs, but significantly increases during neural differentiation. Further, using the recently established in vitro 3D embryogenesis models, ATC measurements reveal that cells continue to stiffen while maintaining pluripotency during epiblast cyst formation. In contrast, during amniotic cyst formation, cells first become stiffer during luminal cavity formation, but softens significantly when cells differentiate to form amniotic cysts. These results suggest that cell stiffness changes not only due to 3D spatial organization, but also with cell fate change. ATC therefore provides a versatile platform for in situ measurement of cellular mechanical property, and cell stiffness may be used as a mechanical biomarker for cell lineage diversification and cell fate specification during embryogenesis.


Assuntos
Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Embrionárias Humanas/metabolismo , Integrinas/química , Microbolhas , Diferenciação Celular/fisiologia , Humanos , Morfogênese/fisiologia , Fenótipo
17.
Biomater Sci ; 6(12): 3139-3151, 2018 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-30307450

RESUMO

Constructs of magnetic nanocomposite hydrogels microencapsulated with stem cells are of great interest as smart materials for tissue engineering and regenerative medicine. Due to the short shelf life of such biocomposites at an ambient temperature, their long-term storage and banking at cryogenic temperatures are essential for the "off-the-shelf" availability of such biocomposites for widespread clinical applications. However, high-quality cryogenic recovery of stem cell-nanocomposite hydrogel constructs has not yet been achieved due to the damage to cells and/or microstructures of hydrogel constructs caused by ice formation, particularly during warming from cryogenic temperatures. Herein, stem cell-magnetic nanocomposite hydrogel constructs, which have an inherent magnetothermal property provided by embedded magnetic nanoparticles, are explored to achieve ultra-rapid cryogenic warming. The binding of water molecules by the hydrogel combined with the magnetothermal heating greatly suppressed ice formation during both cryogenic cooling and warming. Thus, the cryogenic recovery of nanocomposite hydrogel constructs with intact microstructures and fully functional stem cells from ultra-low temperatures was successfully achieved. We further demonstrated that magnetic nanocomposite hydrogels microencapsulated with stem cells could be conveniently manipulated for a self-assembled 3D culture. Together, we have developed a highly efficient and easy-to-perform approach for the cryogenic recovery of stem cell-encapsulated magnetic nanocomposite hydrogel constructs. Our results will facilitate the applications of such stem cell-magnetic nanocomposite hydrogels in regenerative medicine and tissue engineering.


Assuntos
Alginatos/química , Hidrogéis/química , Nanopartículas de Magnetita/química , Nanocompostos/química , Células Cultivadas , Congelamento , Calefação , Humanos , Hidrogéis/efeitos adversos , Hidrogéis/farmacologia , Campos Magnéticos , Células-Tronco Mesenquimais/efeitos dos fármacos
18.
Sci Rep ; 8(1): 12977, 2018 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-30154528

RESUMO

Mechanical forces play critical roles in influencing human embryonic stem cell (hESC) fate. However, it remains largely uncharacterized how local mechanical forces influence hESC behavior in vitro. Here, we used an ultrasound (US) technique, acoustic tweezing cytometry (ATC), to apply targeted cyclic subcellular forces to hESCs via integrin-bound microbubbles (MBs). We found that ATC-mediated cyclic forces applied for 30 min to hESCs near the edge of a colony induced immediate global responses throughout the colony, suggesting the importance of cell-cell connection in the mechanoresponsiveness of hESCs to ATC-applied forces. ATC application generated increased contractile force, enhanced calcium activity, as well as decreased expression of pluripotency transcription factors Oct4 and Nanog, leading to rapid initiation of hESC differentiation and characteristic epithelial-mesenchymal transition (EMT) events that depend on focal adhesion kinase (FAK) activation and cytoskeleton (CSK) tension. These results reveal a unique, rapid mechanoresponsiveness and community behavior of hESCs to integrin-targeted cyclic forces.


Assuntos
Diferenciação Celular , Transição Epitelial-Mesenquimal , Células-Tronco Embrionárias Humanas/metabolismo , Mecanotransdução Celular , Ondas Ultrassônicas , Linhagem Celular , Citoesqueleto/metabolismo , Quinase 1 de Adesão Focal/metabolismo , Células-Tronco Embrionárias Humanas/citologia , Humanos
19.
Biomaterials ; 181: 280-292, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30096562

RESUMO

During normal development, the extracellular matrix (ECM) regulates cell fate mechanically and biochemically. However, the ECM's influence on lineage reprogramming, a process by which a cell's developmental cycle is reversed to attain a progenitor-like cell state followed by subsequent differentiation into a desired cell phenotype, is unknown. Using a material mimetic of the ECM, here we show that ligand identity, ligand density, and substrate modulus modulate indirect cardiac reprogramming efficiency, but were not individually correlated with phenotypic outcomes in a predictive manner. Alternatively, we developed a data-driven model using partial least squares regression to relate short-term cell states, defined by quantitative mechanosensitive responses to different material environments, with long-term changes in phenotype. This model was validated by accurately predicting the reprogramming outcomes on a different material platform. Collectively, these findings suggest a means to rapidly screen candidate biomaterials that support reprogramming with high efficiency, without subjecting cells to the entire reprogramming process.


Assuntos
Materiais Biocompatíveis/farmacologia , Biologia de Sistemas/métodos , Animais , Cálcio/metabolismo , Células Cultivadas , Reprogramação Celular/efeitos dos fármacos , Dimetilpolisiloxanos/química , Matriz Extracelular/química , Mecanotransdução Celular/efeitos dos fármacos , Camundongos
20.
Huan Jing Ke Xue ; 39(6): 2963-2970, 2018 Jun 08.
Artigo em Chinês | MEDLINE | ID: mdl-29965656

RESUMO

The emissions characteristics of 16 kinds of polycyclic aromatic hydrocarbons (PAHs) in ambient air during the waste tire retreading process (open-air storage, mixing, vulcanization, and grinding processes) and in workers' dormitory were analyzed. In addition, the occupational health risk of the workers was evaluated. Results showed that PAHs were detected in all retreading processes and in the workers' dormitory. The highest concentration site was the mixing process, followed by open-air storage and vulcanization process. The lowest concentration point was in the grinding process. The average concentration of PAHs in the workers' dormitory was 11.1 ng·m-3. The PAHs at all sampling points were largely phenanthrene (Phe), fluoranthene (Flu), anthracene (Ant), and pyrene (Pry), which also had a stronger linear correlation with the total PAH concentration. An analysis of the benzene rings showed that three ring and four ring were the majority, while two ring, five ring, and six ring components accounted for less than 10%. Results of the possible influencing factors of the PAHs revealed that the open-air storage and dormitory might be affected by a combustion source, but the mixing, vulcanization, and grinding processes might be affected by rubber oil. The principal component analysis (PCA) and cluster analysis showed that the spatial location of all sites would significantly influence the distribution of PAHs during the tire retreading process. The health risk assessment showed that occupational workers had a lower risk of lifelong cancer, and there was little influence on life expectancy.


Assuntos
Poluentes Atmosféricos/efeitos adversos , Exposição Ocupacional/análise , Hidrocarbonetos Policíclicos Aromáticos/efeitos adversos , Reciclagem , Resíduos Sólidos , Monitoramento Ambiental , Humanos , Medição de Risco
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